The current model for induction of long-term synaptic modifications assumes a transient rise of NMDA receptor-mediated calcium influx into postsynaptic dendritic spines, acting as individual calcium compartments independent of the parent dendrite. It is in this context that simultaneous electrophysiological recording with high-resolution calcium imaging will make fundamental contributions to the issues of long-term synaptic plasticity. A fluorescence microscope for combined patch-clamp electrophysiology and digital imaging has been used to characterize changes in the intracellular concentration of calcium in spines and dendrites of CA1 hippocampal pyramidal neurons maintained in slice culture. High-frequency stimulation of afferent fibers under conditions of voltage-gated channel and ionotropic glutamate receptor blockade leads to activation of NMDA receptor-mediated postsynaptic currents and calcium increases occurring first in spine heads reaching 1.5-2.5 uM. Dendritic segments devoid of nearby spines, 10-50 um away from the activated spines did not show calcium transients. Some activated spines reached steady-state levels around 100-500 ms from the start of the tetanus, while others continued to increase their calcium concentration up to the end of the 1 sec stimulus train. This result suggest the existence of heterogeneity in buffering mechanisms between dendritic spines. Postsynaptic currents and spine calcium transients are completely and reversibly blocked by APV, further indicating the involvement of NMDA receptors in these responses. These results indicate that the combined application of digital microfluorometric imaging of fluorescent indicators sensitive to free calcium with patch clamp recordings is specially suited to study the role of different neuronal compartments in synaptic integration and plasticity by simultaneous, non-invasive access to large regions of the dendritic tree in individual CNS neurons.